The enhancement of IL-12–induced STAT5 and STAT1 activation by IFN-γ is dependent on IL-12. (A) PBMC were cultured with either PHA alone, PHA + IFN-γ, or PHA + IFN-γ + a neutralizing IL-12 antibody (10 μg/mL), and T cells were then stimulated with medium alone (lanes 1, 4, and 7), IL-12 (lanes 2, 5, and 8), or IL-2 (lanes 3, 6, and 9). Western blots were performed with antibodies to phospho-STAT5 and phosphoSTAT1 (upper panels), as well as with antibodies to STAT5 and STAT1 (lower panels). Results are representative of two separate experiments. (B) PBMC were cultured with PHA alone, PHA + IL-12 1 pmol/L, or PHA + IL-12 1 pmol/L + a neutralizing IFN-γ antibody (5 μg/mL). T cells were then stimulated with medium alone (lanes 1, 4, and 7), IL-12 (lanes 2, 5, and 8), or IL-2 (lanes 3, 6, and 9), and Western blots were performed as described in (A). Results are representative of two separate experiments.